A two-photon (TP) absorbing fluorescent substance is excited by receiving energy with a wavelength corresponding to a half of a one-photon (OP) absorbing fluorescent substance to have an excitation characteristic. As a result, the two-photon (TP) absorbing fluorescent substance has advantages of high cell permeability, low cell destruction, and a low quenching effect by a material such as hemoglobin in vivo, and has a characteristic capable of implementing a very high resolution because only a focus portion thereof is excited: (a) Denk, W. et al., Science, 1990, 248, 73; (b) Zipfel, W. R. et al., Nat. Biotechnol, 2003, 21, 1369; (c) Helmchen. F. et al., Nat. Methods, 2005, 2, 932.
Acedan (2-acyl-6-dimethylamino-naphthalene), as a representative two-photon absorbing fluorescent substance, has high photo-stability and a large two-photon absorption cross-section value, and has an advantage of providing a bright image in two-photon microscopy: (a) Kim, H. M. et al., Angew. Chem. Int. Ed, 2007, 46, 3460; (b) Kim, H. M. et al., Angew. Che,. Int. Ed. 2007, 46, 7445; (c) Lee, J. H. et al., J. Am. Chem. Soc. 2010, 132, 1216.
Coumarin is a one-photon absorbing fluorescent substance which is widely used due to high fluorescence efficiency (quantum yield), photo-stability, brightness, and the like. However, a development research of the two-photon absorbing fluorescent substance using coumarin derivatives is scarce up to now.
A method of analyzing or sensing a specific substrate includes electrical, electrochemical, thermal, magnetic, fluorimetric sensing methods, and the like. Among the methods, the fluorimetric sensing method has advantages of high sensitivity and resolution, low price, portability, and the like. Recently, as a research trend of a fluorimetric sensing probe, a reaction-based fluorescent probe accompanying a chemical reaction by responding to a substrate has received attention. The reason is that the reaction-based fluorescent probe is sensitized with the substrate to be selectively analyzed under an ambient environment in which the reaction is irreversible and complicated or an in-vivo condition (Jun, M. E. et al., Chem. Commun. 2011, 47, 7583). Up to now, researches based on the two-photon absorbing fluorescent substance of the reaction-based fluorescent probe are not particularly reported.
A representative method among methods of sensing a substrate such as an enzyme having biological characteristics is a fluorescence sensing method. Accordingly, various kinds of enzyme sensing fluorescent probes have been developed: (a) Takeuchi, M. et al., Acc. Chem. Res. 2001, 34, 865; (b) Peczuh, M. W. et al., Chem. Rev. 2000, 100, 2479. However, the enzyme sensing fluorescent probes are almost based on the one-photon absorbing fluorescent substance: (a) Chen, G. et al., J. Am. Chem. Soc. 2005, 127, 4544; (b) Zhou, W. et al., J. Am. Chem. Soc. 2006, 128, 3122; (c) Alber, A. E. et al., Chem. Commun. 2007, 4647; (d) Aw. J. et al., Chem. Asian. J. 2010, 5, 1317.
A monoamine oxidase (MAO) enzyme is an important factor of maintaining homeostasis of adrenaline, serotonin, dopamine, or the like which is a neurotransmitter. In case of nervous system diseases such as Parkinson's disease and depression, the MAO activity is not controlled and thus the diseases are generated, and in the remedy of the diseases, sensing enzyme activity has an important research value (Chen, G. et al., J. Am. Chem. Soc. 2005, 127, 4544). The MAO enzyme is divided into an MAO A and an MAO B according to a structure and characteristics.